A. Lamure

Thermal analysis characterization of aortic tissues for cardiac valve bioprostheses

Two multistep extractions were achieved on porcine aortic tissues to obtain acellular matrices used for cardiac bioprostheses. The evaluation of structural modifications and the possible damage of extracellular matrix fibrous proteins were investigated by means of thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). Protein-water interactions and degradation temperatures were determined by TGA. DSC was used to characterize protein thermal transitions (glass transition and denaturation), which provided information on the dynamic structure of the aortic tissue components. Sodium dodecyl sulfate (SDS) extraction had a destructuring effect, while Triton and cholate treatments did not affect the structural integrity of either elastin and collagen. A DSC comparison showed that SDS destabilizes the collagen triple helical domain and swells the elastin network.

Dielectric relaxations of collagen and elastin in the dehydrated state

The dielectric properties of collagen and elastin, two major components of connective tissues that greatly differ in their secondary structure, were investigated in the air-dried state and at high temperatures (100–220°C). Two techniques were used to study the dielectric relaxations of both proteins: thermally stimulated currents (TSC), an isochronal spectrometry running at variable temperature, analogous to a low frequency spectroscopy (10−3–10−2 Hz) and dynamic dielectric spectroscopy (DDS), performed isothermally with the frequency varying from 10−2 to 106 Hz. A main relaxation mode is evidenced by the two techniques for collagen and elastin, located at 90°C and 145°C, respectively, at 10−3 Hz. The combination of TSC and DDS experiments and the determination of the activation parameters of the relaxation times give information on the molecular mobility of the proteins, in the glassy state and in the liquid state. Major differences between the relaxation behavior of elastin and collagen have been discussed with the fragility concept of Angell and correlated with the structure of both proteins.

Characterisation of elastin and collagen in aortic bioprostheses

Porcine aortic valves used as cardiac valve bioprostheses are well adapted to physiological functions in the short term, but they lack long-term durability. Several multi-step extractions have been performed to obtain a perfectly acellular matrix. A new physical methodology is proposed to evaluate the resulting fibrous protein damage after biochemical extraction (TRI-COL and SDS). Thermal analysis techniques are adapted to collagen and elastin characterisation in the solid state. The aortic tissue thermal transitions are determined by differential scanning calorimetry (DSC): elastin glass transition is observed around 200°C, and collagen denaturation is observed around 230°C. These parameters are characteristic of the elastin network arrangement and of collagen triple-helix stability. The technique of thermostimulated currents (TSC) is well suited to specify the chain dynamics of proteins. The lowtemperature relaxations observed in both collagen and elastin are associated with localised motions, whereas the high-temperature modes are attributed to more delocalised motions of the chains. Therefore TSC and DSC spectrometries allow physical parameters specific to collagen and elastin to be obtained and their interaction in aortic tissues to be determined. According to the significant evolution of these parameters on SDS samples, the destabilising effect of this detergent is highlighted.

Polarization Phenomena in Collagens from Various Tissues

The various levels of organization of collagens extracted from different tissues (tendon, skin, cartilage) give rise to several polarization phenomena induced by molecular movements of dipolar entities. The corresponding dielectric relaxation time spectrum has been investigated by the Thermally Stimulated Current technique. The contribution of cooperative movements ofpolar and apolar sequences along the main chain has been identified: the corresponding relaxation times are given by Arrhenius equations following a compensation law. The relaxation times obey a Fulcher-Vogel equation and have been assigned to motions delocalized along the whole tropocollagen molecule. Special attention has been paid to dipolar reorientations associated with intra and intermolecular mobility which have been found to be specific of the type of collagen and of its interaction with the other constituents of the organic matrix.

Reorientable Electric Dipoles and Cooperative Phenomena in Human Tooth Enamel

SummaryA preliminary investigation of electric dipole reorientability in human tooth enamel (TE) in comparison to that in hydroxyapatite (OHAp) has been made with the fractional-polarization form of the thermally stimulated currents (TSC) method. The reorientable dipoles are the structural OH− ions. The OHAp exhibited compensation phenomena at 211.5°C and at 356°C which are associated here with the hexagonal form becoming quasi-statically stabilized and dynamically stabilized, respectively, against the monoclinic form. TE specimens were pretreated at various temperatures. All showed the onset of cooperative motions that could quasi-statically stabilize the hexagonal form at the same temperature, approximately 212°C, as did OHAp, even though the TE was already statically stabilized in the hexagonal form. Parts of the TSC spectra that did not conform to the 212°C compensation changed progressively with pretreatment temperature. Loss of incorporated H2O is identified as the most probable cause of most of these changes. This work shows considerable promise for TSC as a tool for further quantitative investigation of TE.

Dielectric characterization of collagen, elastin, and aortic valves in the low temperature range.

The low temperature dielectric relaxation of porcine aortic valves and its main macromolecular proteins, i.e. elastin and collagen, have been investigated in the dry state and at low levels of hydration by thermally stimulated currents spectrometry, with an equivalent frequency of 10-3 Hz. Two secondary relaxation modes, labeled γ and β with increasing temperature, are found for the three materials. Since the γ-mode is independent upon hydration while the β-mode is strongly plasticized by water, these relaxation modes have been attributed to localized motions of the polypeptidic chains containing apolar and polar residues, respectively. The deconvolution of the β-mode by fractional polarization gives the experimental distribution of the dielectric relaxation times of the three materials, and allows us to deduce the activation parameters of each elementary process. These analyses shows the existence of compensation phenomena between the activation parameters, implying cooperative mechanisms. The occurrence of these phenomena with their characteristic parameters are used to specify the origin of the localized relaxation modes in collagen and elastin, and to assign the specific role of each protein in the aortic valves.

Alterations in the chain dynamics of insoluble elastin upon proteolysis by serine elastases

The high temperature dielectric relaxations of purified and elastolized ligamentum nuchae elastin in the dry state have been investigated by thermally stimulated depolarization current spectrometry, with an equivalent frequency comprised between 10(-2) and 10(-3) Hz. A main relaxation mode, located close to 150 degrees C and attributed to the dielectric manifestation of a glass transition, is found for all samples. After decomposition by the fractional polarization method, the analysis of the high temperature mode shows the existence of two relaxation mechanisms: a cooperative one, associated with flexible zones of the protein, and an isoenthalpic one, corresponding to more ordered and constrained zones. The activation parameters of the two mechanisms are dependent on the extent of elastolysis and on the nature of enzyme (pancreatic elastase vs leukocyte elastase). Both enzymes influence the dielectric behavior of elastin in a similar way: the activation enthalpy maximum of the relaxing units located in the flexible zones, characteristic of the cooperative length, decreases with increasing hydrolysis. Moreover, the isoenthalpic mechanism becomes cooperative at the highest extent of elastolysis, which highlights release of constraints in ordered zones. Nevertheless, the differences found between the two enzymatic hydrolyses are characteristic of distinct sites of cleavage in the elastin network.

Thermal analysis of amorphous phase in a pharmaceutical drug

Thermally Stimulated Current (TSC) spectroscopy and Differential Scanning Calorimetry (DSC) have been applied to the characterization of the microstructure of a pharmaceutical drug.The dielectric relaxation spectrum shows two modes located in the temperature range of the glass transition. They have been attributed to the molecular mobility in the “true amorphous phase” and in the ≪rigid amorphous region≫.

Influence of defects and substitutions on polarization phenomena of bioelectrets: apatites

Nonstoichiometric hydroxyapatites and carbonated hydroxyapatites are characterized by the thermally stimulated current technique. Since dielectric energy losses are cooperative, the compensation temperature has been chosen as fingerprint of the local order. Relaxation map analysis shows the existence of either monophasic or biphasic structures and analysis of polarization phenomena gives information on localization and interaction of defects with structural ions of the apatites. >

Polarization phenomena in collagens from various tissues

A comparative study of polarization phenomena in collagens extracted from various tissus (tendon, skin, cartilage) is presented. For this investigation, the ThermoStimulated Current (TSC) technique has been used. A special attention has been paid to dipolar reorientations associated with intra molecular mobility which has been found to be specific of the typical collagen and of its interaction with the other constituants of the organic matrix.